Diabetes affects almost 9% of the United States population and 20-50% of diabetic patients experience painful diabetic neuropathy. Painful diabetic neuropathy (PDN) results in increased somatosensory responsiveness, increased patient-reported pain, and has a greater prevalence in type 2 versus type 1 diabetic patients. Ninety percent of diabetic patients are type 2, pointing to the need to understand the mechanisms that underlie PDN. Plasma methylglyoxal, a catabolic cellular metabolite of glucose, is markedly increased in hyperglycemic diabetic patients, mice, and rats. Very recent studies also suggest that methylglyoxal causes pain via TRPA1-dependent neuronal sensitization, but this mechanism has only been investigated in the streptozotocin (STZ) model of type 1 diabetes. Other chronic studies in the STZ model indicate increased neuronal responsiveness to peripheral stimulation (sensitization) and pain. Lack of comparable longitudinal studies in a rat model of type 2 diabetes is an important gap in knowledge. The proposed studies will use the Zucker Diabetic Fatty (ZDFfa/fa) rat model of type 2 diabetes to test the central hypothesis that hyperglycemia-induced increases in methylglyoxal cause painful diabetic neuropathy in type 2 diabetes. By understanding how changes in peripheral nerves lead to alteration of sensory processing in the spinal cord we can learn how to effectively treat painful diabetic neuropathy. The goal of this project is to discover new therapeutic targets for PDN treatment. Specific Aim 1 will test the hypothesis that methylglyoxal produces spontaneous pain-like behavior in type 2 diabetes. Reactive methylglyoxal directly activates nociceptive sensory neurons to cause pain. We will attempt to inhibit methylglyoxal-induced pathology by several different methods, including the FDA-approved diabetes therapy pioglitazone, and assess whether pain is diminished. Specific Aim 2 will test the hypothesis that methylglyoxal contributes to painful diabetic neuropathy via increasing TRPA1-dependent sensory neuron activity. TRPA1 is a protein expressed in sensory neurons that results in painful sensation upon activation. It is thought that methylglyoxal causes sustained activation of TRPA1, which is at least partially responsible for pain in type 2 diabetes. We will use behavioral assays, mechanistic pharmacology, and calcium imaging as a marker for sensory neuron activation to test the overall hypothesis that type 2 diabetic pain is mediated by glucose-related neurological deficits. Our long-term objective is to substantially reduce the burden of neuropathic pain in type 2 diabetes.